This application claims benefit of foreign priority from Canada patent application 2,267,481, filed Mar. 30, 1999.
The present invention relates to a monoclonal antibody which demonstrates specific binding to human estrogen-responsive isoenzyme of Leucine Aminopeptidase (es-LAPase). The present invention also relates to the hybridoma cell line, designated as 7B6, and the monoclonal antibody produced by the same. The present invention further relates to a diagnostic system using the monoclonal antibody from the hybridoma cell line 7B6, to detect blood, serum or plasma levels of the estrogen responsive isoenzyme of Leucine Aminopeptidase. The antibody is particularly useful for rapid diagnostic tests for breast cancer.
The identification and characterization of risk factors and their molecular implications in the pathophysiology of human diseases such as breast cancer is essential for designing efficient diagnostic assays and therapeutic compounds. Amongst the various risk factors associated with the onset of early events leading to Breast Cancer, estrogen and estrogen-like compounds with estrogenic mimicking activity remain the most important determinants in the early events and progression of breast carcinogenesis. Under normal physiological conditions, there are several tissues whereby estrogenic steroids have been shown to play a critical function. These include the development of the reproductive tract, particularly secondary organs, such as the mammary glands. In addition, estrogens are also involved in the fine regulation of bone growth, liver and cardiovascular function and the estrus cycle, most likely through the induction of cell proliferation in target tissues [Sutherland R. L. et al., pp. 197-215, Elsevier Science Publishing B. V., Amsterdam., Shekhar P. V. M., et al., J. NatL Cancer. Inst., 89: 1774-1782.]. The response of such a variety of tissues to estrogen stimulation can explain in part its active role in the development and progression of different human carcinomas and in particular of Breast Cancer. Although the precise molecular mechanisms by which estrogen stimulation regulates various physiological functions requires further elucidation, this steroid is involved in both xe2x80x9cimmediate-earlyxe2x80x9d and xe2x80x9cearlyxe2x80x9d events of cell function. In this regard, it appears that immediate early events induced by estrogen lead to an increased cellular proliferation most likely through the reduction in the cell cycle by accelerating the rate at which cells progress from the Gl phase towards the S phase. Recently, it has been proposed that estrogen promotes cellular proliferation by co-activating at similar estrogen concentrations, the expression of cyclin Dl-Cdk4 and cyclin E-Cdk2, two critical and potentially interrelated Gl regulatory peptides [Prall, O. W. J., et al., J. Biol. Chem., 272: 10882-10894.].
Diagnostic assays are available for breast cancer. For example, imaging techniques such as ultrasounds and x-rays (mammographies) are widely used to detect tumours. These imaging techniques, however, suffer from a limitation in the resolution of the image which prevents the detection of tumours below a certain size.
Histological analysis of biopsies is also a common procedure for the diagnosis of breast cancer and this technique relies on identification of visible phenotypes of the cells. However, this analysis is somewhat subjective and depends on the skill of the examiner.
Molecular diagnostics assays are also available. Among the most widely relied upon is an assay which measures estrogen and progesterone receptors on cells obtained from biopsies. This assay is useful to determine whether a particular type of cancer will be responsive to hormonal therapy.
Flow cytometry can also be used to measure parameters, such as DNA content and the proportion of cells in a particular phase of the cell cycle, that correlates with the presence of cancerous cells.
However, histology, estrogen and progesterone receptor analysis and flow cytometry all require that biopsies be taken and that the sample be extensively processed.
Therefore, there is a need for more rapid and less invasive methods of diagnosing breast cancer. In particular, sensitive assays to detect the presence of cellular markers, preferably in the blood, which reflect the presence of metastasis are desirable. Even more desirable, given the prominent role of estrogen in the progression of breast cancer, are cellular markers, other than estrogen and progesterone receptors, responsive to estrogen.
The present invention relates to a monoclonal antibody which demonstrates specific binding to human estrogen-stimulated leucine aminopeptidase (es-LAPase). The present invention further relates to a diagnostic system using the monoclonal antibody to detect blood, serum, plasma or tissue levels of the es-LAPase.
Thus, according to the present invention there is provided a monoclonal antibody which is specific for es-LAPase.
In a further embodiment of the present invention there is provided a method of detecting breast cancer in a patient by determining the level of es-LAPase in a sample.
This invention is also directed to a method for detecting a metastatic cancer in a patient by determining the level of es-LAPase in a sample.